Direct Visualization of Structural Maintenance of Chromosomes by Protein-Mediated Loop Extrusion Using High-Speed Atomic Force Microscopy

Project Overview

A strand of DNA nearly meters long is folded into a small cellular nucleus 10 µm in diameter by a loop extrusion process. This process is organized by Structural Maintenance of Chromosomes (SMC) Protein, which is a large ATPase in all kingdoms of life. However, the mechanism of SMC and DNA interaction is still not well understood. In the current project, we utilized high-speed atomic force microscopy, one of the cutting-edge techniques, to characterize the SMC and DNA interaction in real time. We provided direct evidence for the hypothesis that Bacillus subtilis SMC (BsSMC) proteins serve as ATP motors extruding DNA strands. The pre-stretched DNA on the mica surface was taken as a scaffold in this study. Our results demonstrated that SMC proteins could adopt multiple configurations, including O-shaped, I-shaped, and transient dimers with Adenosine 5'-triphosphate (ATP) addition. As a comparison, the coarse-grained Molecular Dynamics (MD) simulation was performed to interpret the dimerization of SMC. Such SMC dimers bind to DNA molecules through either head and hinge domains and form the loop structure gradually. In addition, the dimer is quite dynamic and changes its conformation over time.

Mission Impact

Our results shed light on the interaction of SMC and DNA molecules, help to verify the loop-extrusion hypothesis, establish the SMC proteins as ATP-driven processive motors, and significantly advance our understanding of DNA packaging in chromosomes. Current data obtained in the LDRD project advances LLNL's core competency in bioscience and bioengineering, and will be used for the NIH (R21) proposal.